The scientific community has long used methods of descriptive observation as a means of reducing the mysteries of nature into discrete and finite systems. Analogous to the utility of telescopes in describing spectral features of the night sky, optical microscopy seeks to mitigate the limitations of the human eye when investigating the endogenous microcosms all around us. The key advantage inherent in these instruments is the improved resolution afforded by the magnification and image contrast of a sample. Optical microscopy has found applications in a broad spectrum of disciplines where surface characterization or the determination of structural morphology is needed. Population screening and clinical diagnostics by optical microscopy are possible thanks to recent advances in histology (i.e. study of tissue). The advantage of optical microscopy versus other methods of analysis is the relatively rapid, low cost, low maintenance platform of testing inherent in routine screening methods.
The sample preparation template can be executed in eight steps: 1) preserve specimen to prevent structural degradation using a formaldehyde based reagent 2) embed the sample in a paraffin wax for structural support 3) cut a thin slice using a specialized device that delivers a sample thickness on the order of microns (microtome) 4) remove wax with solvents to recover the native specimen 5) optimize color contrast using specific stains. The goal of this step is to optimize the sample image by adjusting the colors of the individual components (organelles) of the cells that make of up the tissue structure. There are four basic types of tissue: muscle tissue, nervous tissue, connective tissue, and epithelial tissue. It follows that optimal sample preparation procedures are dependant on the nature of the sample (e.g. selection of dyes etc.) A pedestrian visualization of this process can be described as the process of adjusting the color tones of a television unit (e.g. light, dark, contrast, brightness). This step improves the overall resolution of the method 6) image the sample under microscope by selecting the strength of magnification and light sources 7) compare against standards or previously reported test specifications. Medical professionals will often compare a sample from a patient to control-healthy and control-diseased tissue samples in clinical diagnostics. 8) report results.
Note each step of the preparation procedure may induce artifacts in the image that may have a deleterious effect on the interpretation of results. It follows that a number of strategies over time have been evaluated to minimize the number of steps in the sample preparation procedure. Cold temperature techniques are currently in vogue and practiced regularly by hospitals and medical research laboratories. In the template above, steps 2-4 reflecting the wax application can be circumvented using a cold approach, freeze the sample and cut. Commercial available devices are available for these procedures.
